Mitochondria are the site of the essential process of respiration in eukaryotic cells, and contain DNA separate from the nucleus. The mitochondrial genome encodes only a few of the proteins needed for mitochondrial function, with the remainder imported from the cytoplasm from nuclear-encoded genes. Mitochondrial sequence mutations and deletions have been linked with various human diseases, including cardiomyopathy, neuropathy and various aging disorders. In plants, mitochondrial genome alterations are associated with generation of some novel open reading frames and cytoplasmic male sterility, which results in abortive pollen production. The overall goal of this research is to determine whether mtDNA recombination is essential for genome replication or maintenance, or whether it is involved in repair of mtDNA damage in higher plants. We are using Arabidopsis thaliana as a model system for these studies, as the complete genome (nuclear, mitochondrial and chloroplast) is available, and propagation and genetic studies with this plant are relatively easy. The goals of this renewal proposal are to: 1. Examine the effect of alteration of expression of the Arabidopsis recA gene on plant growth and mitochondrial ultrastructure, function, and genome structure; 2. Complete characterization of the two Arabidopsis thaliana mitochondria-targeted homologues of the E. coli ssb gene; 3. Determine targeting and function of the Arabidopsis homologue of yeast Pif1 helicase, and analyze the effect of alteration of expression. The effect of overexpression of the recA, ssb, helicase and other yet to be identified genes involved in DNA recombination, driven either by global or tissue-specific strong promoters, will be determined in transgenic plants. Disruption or reduced expression of endogenous genes by gene knockout or RNAi technology will also be examined to determine if mitochondrial DNA recombination is essential for proper growth and development, or if gene disruption leads to greater susceptibility to DNA damaging agents. Our findings will provide greater insights into processes affecting mitochondrial genome integrity, which may be extended to mitochondria from other organisms.